The goal of this proposal is to lay the foundation for a full development effort of a synthetic peptide-based multi-flavivirus vaccine. In our previous work on flaviviruses, our group produced a recombinant monoclonal antibody against West Nile virus (WNV) envelope (E) protein using a phage display human library screen. Unexpectedly, the antibody recognized the fusion loop, a flavivirus E protein epitope that mediates viral entry by membrane fusion and that does not normally elicit an antibody response following infection with WN virus or immunization with E protein. The mAb was protective against WN virus infection in mice and could also clear an ongoing WN infection. Remarkably, the mAb also reacted with several other flaviviruses, such as dengue virus, suggesting it may be broadly cross-protective (1). Here we propose that a recombinant fusion loop peptide vaccine that targets the epitope recognized by the mAb can prevent and treat a broad spectrum of flavivirus infections, with WN virus serving as our initial model infection. Unfortunately, Peptides representing this epitope are not highly immunogenic in mice. We therefore propose to synthesize a set of improved fusion loop peptides, modified to increase the immunogenicity of the native peptides. A set of peptides including, multiple antigenic peptide modifications will be used to test this approach using established adjuvant and delivery techniques. We will then challenge mice and evaluate humoral and cellular responses. Candidate formulations that elicit a response will then be evaluated for their neutralizing capacity of WNV. The main goal of this proposal is then to evaluate these candidate formulations against a broad array of Flaviviruses such as WN virus, Kunjin, Japanese encephalitis virus, Murray valley encephalitis, St. Louis encephalitis virus (SLEV), Dengue virus (serotypes 1 to 4), and Yellow fever virus. Finally, any formulations that do show both specific activity against WNV and against a broader set of flaviviruses will be tested in vivo against our well established C3H/HeN mouse Model of neuroinvasive WNV. This pilot approach to a broad based peptide vaccine strategy could have profound impact against multiple flavivirus pathogens including flaviviruses for which no vaccines are currently available. PUBLIC HEALTH RELEVANCE: Human pathogens in the Flavivirus genus cause significant morbidity and mortality worldwide. This project is designed to develop a new vaccine candidate that might protect humans against a variety of Flavivirus infections.